|Source||An E.coli strain, that carries the cloned gene EcoRV from Escherichia coli|
|Assayed on||Lambda DNA|
|Unit definition||One unit of the enzyme is the amount required to hydrolyze 1 μg of Lambda DNA in 1 hour at 37°C in a total reaction volume of 50 μl.|
|Optimal SE-buffer||W (10 mM Tris-HCl (pH 8.5 at 25°C); 10 mM MgCl2; 100 mM NaCl; 1 mM DTT.) + BSA|
|Enzyme activity (%)|
|0 - 10||25 - 50||50 - 75||100||25 - 50||50|
|Storage conditions||10 mM Tris-HCl (pH 7.5); 50 mM NaCl; 0,1 mM EDTA; 1 mM DTT; 200 μg/ml BSA; 50% glycerol; Store at -20°C.|
|Ligation||After 20-fold overdigestion with enzyme 90% of the DNA fragments can be ligated and recut.|
|Non-specific hydrolisis||No nonspecific activity was detected after incubation of 1 μg of Lambda DNA with 20 u.a. of enzyme fro 16 hours at 37°C.
|Reagents Supplied with Enzyme||
10 X SE-buffer W, BSA (except E059T and E060T).|
For E059T and E060T an additional 10X SE-buffer ROSE is supplied.
|Methylation sensitivity||not tested|
|Inactivation 20 minutes under||80oC|
|Notes||High enzyme concentration results in star activity. |
Do not use BSA for long incubation
To obtain 100% activity, BSA should be added to the 1 x reaction mix to a final concentration of 100 μg/ml.
Turbo EcoR V can be used for short time (10 min) DNA digestion as well as for standard reaction. The reaction can be performed using optimal or universal (ROSE) Buffer.
Turbo DNA Digestion:
-Fast DNA analysis
-Fast preparation of vectors for cloning
1 μl of Turbo EcoR V cuts 1 μg of DNA in 1 x SE-Buffer W or universal 1 x SE-Buffer ROSE in 10 min (see the protocol below). A short time of DNA digestion requires high quality purification of DNA sample. The enzyme may be used in standard conditions of DNA cleavage (E059).
Turbo reaction protocol:
20 μl of the reaction volume:
10 x SE Buffer ROSE - 2 μl
DNA - 0,2-1 μg
Nuclease-free water - to 20 μl
+ 1μl of Turbo EcoR V
Incubate at 37°C for 10 min.
|Quality control||Restriction Endonucleases: Quality Control|
|MSDS:||Download MSDS as PDF|
|References:||Kholmina, .V., Rebentish, B.A., Skoblov, Yu.S., et.al. Dokl.Akad. Nauk SSSR 253: 495-497 (1980).|
Chernukhin V.A, Abdurashitov M.A., Tomilov V.N., Gonchar D.A., Degtyarev S.Kh. Comparative analysis of mouse chromosomal DNA digestion with restriction endonucleases in vitro and in silico // "Ovchinnikov bulletin of biotechnology and physical and chemical biology" V.3, No 4, pp 19-27, 2007
|Theoretical diagrams of DNA digestion by this enzyme for the most known DNA substrates:|
To view the fragments length values please point mouse cursor over diagram
M - ladder, 1 - Adeno-2 DNA, 2 - Lambda DNA, 3 - T7 DNA, 4 - pUC19, 5 - pBR322